Single nucleotide polymorphism analysis by allele-specific primer extension with real-time bioluminescence detection in a microfluidic device

A microfluidic approach for rapid bioluminescent real-time detection of single nucleotide polymorphism (SNP) is presented. The method is based on single-step primer extension using pyrosequencing chemistry to monitor nucleotide incorporations in real-time. The method takes advantage of the fact that the reaction kinetics differ between matched and mismatched primer-template configurations. We show here that monitoring the initial reaction in real time accurately scores SNPs by comparing the initial reaction kinetics between matched and mismatched configurations. Thus, no additional treatment is required to improve the sequence specificity of the extension, which has been the case for many allele-specific extension assays. The microfluidic approach was evaluated using four SNPs. Three of the SNPs included primer-template configurations that have been previously reported to be difficult to resolve by allele-specific primer extension. All SNPs investigated were successfully scored. Using the microfluidic device, the volume for the bioluminescent assay was reduced dramatically, thus offering a cost-effective and fast SNP analysis method.     

Ternary and binary copper(II) complexes: synthesis,characterization, ROS-inductive,proteasome inhibitory,and anticancer properties

Three ternary copper(II) complexes, [Cu(phen)(L-phe)Cl]·2H₂O, [Cu(phen)(L-leu)Cl]·4½H₂O, and [Cu(phen)(L-tyr)Cl]·3H₂O, and four binary copper(II) complexes, [Cu(phen)Cl₂]_, Cu(L-phe)₂·½H₂O, Cu(L-leu)₂·½H₂O, and Cu(L-tyr)₂·H₂O (where phen = 110-phenanthroline, L-phe = L-phenylalanine, L-tyr = L-tyrosine, L-leu = L-leucine and Cl^- = chloride), were synthesized and characterized by elemental analysis, spectroscopic techniques (FTIR, UV–visible, fluorescence spectroscopy), magnetic susceptibility, molar conductivity, and lipophilicity measurement. X-ray diffraction determination of a single crystal of [Cu(phen)(L-tyr)Cl] showed two independent molecules in the asymmetric unit, each with the same distorted square pyramidal geometry about copper(II). p-Nitrosodimethylaniline assay revealed that the three ternary complexes were better inducers of reactive oxygen species over time than binary complexes, CuCl₂, and free ligands. All the copper(II) complexes in this series inhibited the three proteolytic activities in the order Trypsin-like > Caspase-like > Chymotrypsin-like. In terms of anticancer properties, the copper(II)-phen complexes had GI50 values of less than 4 μM against MCF-7, HepG2, CNE1 and A549 cancer cell lines, more potent than cisplatin.     

Spectrophotometric Quantitation of Fluoxetine Hydrochloride Using Benzoyl Peroxide and Potassium Iodide

 Fluoxetine hydrochloride reacts with benzoyl peroxide and potassium iodide, after heating for 1 min at 30 °C, to give a blue colour having maximum absorbance at 570 nm. The reaction is selective for fluoxetine with 0.01 mg/mL as visual limit of quantitation and provides a basis for a new spectrophotometric determination. The colour reaction obeys Beer’s law from 0.1 mg/10 mL to 2.0 mg/10 mL of fluoxetine and the relative standard deviation is 0.68%. The qualitative assessment of tolerable amounts of other drugs is also studied. Received September 21, 1998. Revision September 10, 1999.     

A Multi‐Addressable Switch Based on the Dimethyldihydropyrene Photochrome with Remarkable Proton‐Triggered Photo‐opening Efficiency

A series of photochromic derivatives based on the trans‐10b,10c‐dimethyl‐10b,10c‐dihydropyrene (DHP, “closed form”) skeleton has been synthesized and their photoisomerization leading to the corresponding cyclophanediene (CPD, “open form”) isomers has been investigated by UV/Vis and ¹H NMR spectroscopies. Substitution of the DHP core with electron‐withdrawing pyridinium groups was found to have major effects on the photoisomerization efficiency, the most remarkable examples being to enhance the quantum yield of the opening reaction and to allow fast and quantitative conversions at much lower radiant energies. This effect was rationalized by theoretical calculations. We also show that the reverse reaction, that is, going from the open form to the closed form, can be electrochemically triggered by oxidation of the CPD unit and that the photo‐opening properties of pyridine‐substituted DHPs can be efficiently tuned by protonation, the system behaving as a multi‐addressable molecular switch. These multi‐addressable photochromes show promise for the development of responsive materials.     

Selectivity in the addition reactions of organometallic reagents to aziridine-2-carboxaldehydes: the effects of protecting groups and substitution patterns

Good to excellent stereoselectivity has been found in the addition reactions of Grignard and organozinc reagents to N-protected aziridine-2-carboxaldehydes. Specifically, high syn selectivity was obtained with benzyl-protected cis, tert-butyloxycarbonyl-protected trans, and tosyl-protected 2,3-disubstituted aziridine-2-carboxaldehydes. Furthermore, rate and selectivity effects of ring substituents, temperature, solvent, and Lewis acid and base modifiers were studied. The diastereomeric preference of addition is dominated by the substrate aziridines' substitution pattern and especially the electronic character and conformational preferences of the nitrogen protecting groups. To help rationalize the observed stereochemical outcomes, conformational and electronic structural analyses of a series of model systems representing the various substitution patterns have been explored by density functional calculations at the B3LYP/6-31G* level of theory with the SM8 solvation model to account for solvent effects.     

Compact passively q-switched Nd:YAG laser for 2D micromachining

We present a compact passively q-switched diode end pumped Nd:YAG laser at 1064 nm for 2D micromachining. It consists of a 5.5 cm long plano-concave end pumped resonator carrying a Cr:YAG passive q-switch inside the cavity. With an optical conversion efficiency of 46 and 33% the laser emits 1.4 W in CW and 986 mW in q-switched mode at a current of 2.5 A. After using a 2 mm circular aperture the output is seen in TEM₀₀ mode with a single pulse energy of 5 mJ. The laser produced circular holes of diameter 75 μm in 25 μm thick Tantalum foils. Actual results of 1D and 2D machining are shown along with the diffraction patters of the samples.     

Water-Soluble Polyelectrolytes with Ability to Remove Arsenic

Arsenic species can be removed from aqueous solutions using the liquid-phase polymer-based retention, LPR, technique. The LPR technique removes ionic species by functional groups of water-soluble polyelectrolytes (WSP) and then using a ultrafiltration membrane that does not let them pass through the membrane, thus separating them from the solution. The ability of WSP with groups (R)₄N⁺X⁻ to remove arsenate ions using LPR was studied. The interaction and arsenate anion retention capacity depended on: pH, the quaternary ammonium group's counter ion, and the ratio polymer: As(V), using different concentrations of As(V). Water-soluble polychelates were also used for one-step retention of As(III) in solution. The complex of poly(acrylic acid)-Sn, 10 and 20 wt-% of metal gave a high retention of As(III) species at pH 8, although the molar ratio polychelate: As(III) was 400:1. The enrichment method was used to determine the maximum retention capacity (C) for arsenate anions in aqueous solutions at pH 8. In similar conditions, the values of C were 142 mg g⁻¹ for P(ClAETA) and 75 mg g⁻¹ for P(SAETA). The combined treatment of arsenic aqueous solutions by electrocatalytic oxidation (EO) to convert the species of As(III) to As(V) with the LPR technique quantitatively removed arsenic.